CRISPR Screening in Primary Human B Cells to Improve B Cell Therapies

Abstract

Pooled CRISPR screening in primary human B cells has been limited by inefficient lentiviraltransduction and high vector copy numbers that compromise single-cell resolution genetic perturbation. This study established a robust CRISPR screening platform by integrating CD20- targeted Nipah pseudotyped lentiviral vectors for single-copy sgRNA delivery with recombinant Cas9 protein electroporation, achieving efficient transduction while maintaining single vector copy number per cell. Essential gene screens validated the platform's technical feasibility, and application to an epigenetic regulator screen revealed that the RARA-NCOR2-TBL1XR1 repressor complex negatively regulates plasma cell differentiation, with disruption of individual components promoting expression of plasma cell surface markers CD38 and CD138. Furthermore, CRISPR screening for natural killer (NK) cell mediated killing revealed HLA-E as an important factor in preventing B cell lysis. This versatile CRISPR screening platform enables systematic investigation of genetic regulators controlling B cell differentiation, survival, and NK tolerance, with applications spanning fundamental immunology and translational cell therapy development.